Monitoring Human Milk β-Casein Phosphorylation and O-Glycosylation Over Lactation Reveals Distinct Differences between the Proteome and Endogenous Peptidome

Human milk is a vital biofluid containing a myriad of molecular components to ensure an infant's best start at a healthy life. One key component of human milk is beta-casein, a protein which is not only a structural constituent of casein micelles but also a source of bioactive, often antimicrobial, peptides contributing to milk's endogenous peptidome. Importantly, post-translational modifications (PTMs) like phosphorylation and glycosylation typically affect the function of proteins and peptides; however, here our understanding of beta-casein is critically limited. To uncover the scope of proteoforms and endogenous peptidoforms we utilized mass spectrometry (LC-MS/MS) to achieve in-depth longitudinal profiling of beta-casein from human milk, studying two donors across 16 weeks of lactation. We not only observed changes in beta-casein's known protein and endogenous peptide phosphorylation, but also in previously unexplored O-glycosylation. This newly discovered PTM of beta-casein may be important as it resides on known beta-casein-derived antimicrobial peptide sequences.

Automated summary

Human milk contains beta-casein, a protein that undergoes post-translational modifications such as phosphorylation and glycosylation. A study observed changes in phosphorylation and previously unexplored O-glycosylation of beta-casein in two donors over 16 weeks of lactation, indicating the importance of these modifications on antimicrobial peptides derived from beta-casein.